Kobi Benenson supervisor: Ehud Shapiro, Dept of Computer Science & Applied Math Acknowledgements:

1. A nanoscale programmable computing machine with input, output, software and hardware made of biomolecules • Nature 414, 430-434 (2001) Kobi Benenson supervisor: Ehud Shapiro, Dept of Computer Science & Applied Math Acknowledgements: Ehud Keinan (Technion), Zvi Livneh (WIS), Tami Paz-Elizur (WIS), Rivka Adar (WIS), Aviv Regev (WIS), Irith Sagi (WIS), Ada Yonath (WIS)

2. “Medicine in 2050: Doctor in a Cell” Molecular Output Molecular Input Programmable Computer

3. Research goal: Design a simplest non-trivial molecular computing machine (two-state two-symbol finite automaton) that works on engineered inputs

4. Finite automaton: an example An even number of b’s b S0, a S0 S0, b  S1 S1, a  S1 S1, b  S0 a a S0 S1 b Two-states, two-symbols automaton

5. Automaton 1 An even number of b’s S0, a S0 S0, b  S1 S1, a  S1 S1, b  S0 b a b S0

6. Automaton 1 An even number of b’s S0, b  S1 S0, a S0 S0, b  S1 S1, a  S1 S1, b  S0 b a b S0

7. Automaton 1 An even number of b’s S0, a S0 S0, b  S1 S1, a  S1 S1, b  S0 a b S1

8. Automaton 1 An even number of b’s S1, a  S1 S0, a S0 S0, b  S1 S1, a  S1 S1, b  S0 a b S1

9. Automaton 1 An even number of b’s S0, a S0 S0, b  S1 S1, a  S1 S1, b  S0 b S1

10. Automaton 1 An even number of b’s S1, b  S0 S0, a S0 S0, b  S1 S1, a  S1 S1, b  S0 b S1

11. Automaton 1 An even number of b’s S0, a S0 S0, b  S1 S1, a  S1 S1, b  S0 S0 The output

12. Rationale for the molecular design

13. Rationale for the molecular design CTGGCT GACCGA CGCAGC GCGTCG a b

14. Rationale for the molecular design CTGGCT GACCGA CGCAGC GCGTCG a b S0, a S0, b GGCT CAGC

15. Rationale for the molecular design CTGGCT GACCGA CGCAGC GCGTCG a b S0, a S0, b GGCT CAGC S1, a S1, b CTGGCT GA CGCAGC CG

16. Rationale for the molecular design Transitions S0, b CAGCCTGGCTCGCAGCTGTCGC GACCGAGCGTCGACAGCG a b t

17. Rationale for the molecular design Transitions S0, b CAGCCTGGCTCGCAGCTGTCGC GACCGAGCGTCGACAGCG a b t S0, b  S1

18. Rationale for the molecular design Transitions S1, a CTGGCTCGCAGCTGTCGC GAGCGTCGACAGCG b t S0, b  S1

19. Rationale for the molecular design Transitions S1, a CTGGCTCGCAGCTGTCGC GAGCGTCGACAGCG b t S1, a  S1

20. Rationale for the molecular design Transitions S1, b CGCAGCTGTCGC CGACAGCG t S1, a  S1

21. Rationale for the molecular design Transitions S1, b CGCAGCTGTCGC CGACAGCG t S1, b  S0

22. Rationale for the molecular design Transitions S0, t TCGC S1, b  S0

23. Rationale for the molecular design Transitions S0, t TCGC Output: S0

24. Rationale for the molecular design Transition procedure: a concept S0, b CAGCCTGGCTCGCAGCTGTCGC GACCGAGCGTCGACAGCG a b t

25. 4 nt GTCG 8 nt Rationale for the molecular design Transition procedure: a concept S0, b CAGCCTGGCTCGCAGCTGTCGC GACCGAGCGTCGACAGCG a b t S0, b -> S1

26. 4 nt GTCG 8 nt Rationale for the molecular design Transition procedure: a concept CAGCCTGGCTCGCAGCTGTCGC GACCGAGCGTCGACAGCG b t S0, b -> S1

27. Rationale for the molecular design Transition procedure: a concept S1, a CTGGCTCGCAGCTGTCGC GAGCGTCGACAGCG b t S0, b -> S1

28. Rationale for the molecular design Transition procedure: a concept S1, a CTGGCTCGCAGCTGTCGC GAGCGTCGACAGCG b t S1, a -> S1

29. 6 nt GACC 10 nt Rationale for the molecular design Transition procedure: a concept S1, a CTGGCTCGCAGCTGTCGC GAGCGTCGACAGCG b t S1, a -> S1

30. 6 nt GACC 10 nt Rationale for the molecular design Transition procedure: a concept CTGGCTCGCAGCTGTCGC GAGCGTCGACAGCG t S1, a -> S1

31. Rationale for the molecular design Transition procedure: a concept S1, b CGCAGCTGTCGC CGACAGCG t S1, a -> S1

32. 8 nt GCGT 12 nt Rationale for the molecular design Transition procedure: a concept S1, b CGCAGCTGTCGC CGACAGCG t S1, b -> S0

33. 8 nt GCGT 12 nt Rationale for the molecular design Transition procedure: a concept CGCAGCTGTCGC CGACAGCG S1, b -> S0

34. Rationale for the molecular design Transition procedure: a concept S0, t TCGC Output: S0

35. Rationale for the molecular design In situ detection S0, t Detection molecule for S0 output TCGC AGCG Output: S0

36. AGCG Rationale for the molecular design In situ detection Reporter molecule for S0 output TCGC Output: S0

37. 4 nt GTCG 8 nt Inside the transition molecule S0,b -> S1

38. Inside the transition molecule FokI 4 nt GGATGACGAC CCTACTGCTG GTCG 8 nt S0,b -> S1

39. Inside the transition molecule FokI 9 nt 4 nt GGATGACGAC CCTACTGCTG GTCG 8 nt 13 nt S0,b -> S1

40. 9 nt GGATGACGAC CCTACTGCTG GTCG 13 nt Inside the transition molecule FokI S0,b -> S1

42. 6 nt GACC 10 nt Inside the transition molecule S1,a -> S1

43. Inside the transition molecule FokI 9 nt 6 nt GGATGACG CCTACTGC GACC 10 nt 13 nt S1,a -> S1

44. 9 nt GGATGACG CCTACTGC GACC 13 nt Inside the transition molecule FokI S1,a -> S1

45. Inside the transition molecule 8 nt GCGT 12 nt S1,b -> S0

46. Inside the transition molecule FokI 9 nt 8 nt GGATGG CCTACC GCGT 12 nt 13 nt S1,b -> S0

47. 9 nt GGATGG CCTACC GCGT 13 nt Inside the transition molecule FokI S1,b -> S0

48. Inside the transition molecule GGATGACGAC CCTACTGCTG S0 -> S1 GTCG S0 -> S0 GGATGACG CCTACTGC GACC S1 -> S1 GGATGG CCTACC S1 -> S0 GCGT

49. Transition rules: complete list

50. Automata programs used to test the molecular implementation